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Direct comparison by limiting dilution analysis of long-term culture-
initiating cells in human bone marrow, umbilical cord blood, and blood stem
cells
R Pettengell, T Luft, R Henschler, JM Hows, TM Dexter, D Ryder and NG Testa
CRC Department of Experimental Haematology, Paterson Institute for Cancer
Research, Manchester, UK.
Limiting-dilution analysis of long-term culture-initiating cells (LTCIC) is
a quantitative method of estimating hematopoietic stem cell activity in
clinical samples. We compared the numbers of LTCIC in bone marrow (BM),
umbilical cord blood, and blood progenitor cells (obtained from patients
with solid tumors at leukapheresis after mobilization with induction
chemotherapy and filgrastim administration), using a two- stage long-term
culture system and a limiting-dilution technique, scoring cobblestone areas
of greater than 15 hematopoietic cells weekly for up to 8 weeks. Samples
were obtained from 30 normal BMs, 20 human umbilical cords, and 32
leukapheresis products. Direct comparison of LTCIC in the three sources
showed that the median proportions of cells generating hematopoietic foci
from unfractionated mononuclear cells at 5 and 8 weeks, respectively, were
1:13,314 and 1:33,949 for BM, 1:12,506 and 1:34,546 for umbilical cord
blood, and 1:10,302 and 1:12,891 for leukapheresis product. The estimated
proportions of LTCIC from unfractionated mononuclear cells and CD34+ cells
were similar in experiments with leukapheresis product. Leukapheresis
product was superior to umbilical cord blood and cord blood to BM at 5 and
8 weeks of culture (P = .01). In two-stage long-term cultures, more
colonies per flask and CD34+ cells were found in assays of leukapheresis
product than in BM or umbilical cord blood cultures (P = .0005). Results
obtained by this simplified limiting-dilution analysis correlated well with
standard long-term cultures and can be used as a measure of the stem cell
population. These data suggest that the incidence of putative stem cells in
leukapheresis product and umbilical cord blood are at least comparable with
that of BM.
Volume 84,
Issue 11,
pp. 3653-3659,
12/01/1994
Copyright © 1994 by The American Society of Hematology
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